Notwithstanding intensive research for a vaccine in the 12 years since the initial discovery of HIV as the Acquired Immunodeficiency Syndrome (AIDS) virus and 10 years since the molecular cloning and characterization of the AIDS virus, major obstacles remain for HIV vaccine and immunotherapy development. These hurdles include HIV-1 variability, multiple routes/modes of virus transmission, and a lack of understanding of the immune responses necessary for prevention of HIV infection. In an article published on Jul. 28, 1995 (Cell 82:175-176), David Baltimore asked all scientists in the field to take a step back and reflect on why this virus infection, against which 10% of the budget of the U.S. National Institutes of Health (NIH) is deployed, remains so enigmatic.
There was early optimism for efficacious recombinant HIV-1 envelope subunit vaccines (e.g., gp120 and gp160 vaccine products) given that vaccinee sera from several clinical trials were capable of neutralizing laboratory isolates of HIV-1 in vitro (Belshe et al., JAMA, 1994, 272:475; Keefer et al., AIDS Res Hum Retroviruses, 1994, 10:1713). This optimism was shaken when the vaccinee sera were found to be largely ineffective in neutralizing HIV-1 primary patient isolates (Hanson, AIDS Res Hum Retroviruses, 1994, 10:645; Mascola et al., J Infect Dis, 1996, 173:340). These disappointing findings led NIH to decide in June 1994 to postpone costly large-scale efficacy trials of several recombinant envelope protein based HIV subunit vaccines.
Primary isolates of HIV-1 are obtained by limited cultivation of patient peripheral blood mononuclear cells (PBMCs) or plasma with uninfected PBMCs. They closely resemble HIV strains responsible for human infection in the field (Sawyer et al., J Virol, 1994, 68:1342; Cornelissen et al., J Virol), 1995, 69:1810). Primary isolates can be readily distinguished from the commonly used laboratory-adapted T-tropic viruses such as IIIb/LAI, SF2, and MN, which have been passaged over time in human T-lymphoid cell lines and are well-adapted to grow in these T cell lines. First, most primary isolates are M-tropic. They do not readily grow in cultured T cell lines, rather, they are monocytes-or macrophage-tropic, although they can also infect primary T cells (Cheng-Mayer et al., Science, 1988, 240:80). Second, primary isolates are highly resistant to in vitro neutralization by recombinant soluble forms of the viral receptor protein CD4 (rsCD4) requiring 200-2700 times more rsCD4 than laboratory strains for comparable neutralization (Daar et al., PNAS USA, 1990, 87:6574-6578). Third, primary isolates are also resistant to neutralizing antibodies elicited by the use of gp120 vaccines (Mascola et al.).
Primary isolates include both syncytium inducing isolates (SI) that induce syncytium formation in PBMC culture and non-syncytium-inducing (NSI) isolates. Among the SI primary isolates, most will replicate in the especially HIV-sensitive T cell line MT2, but few can replicate in the less permissive transformed T cell lines such as CEM or H9 that are commonly used for the culture of laboratory-adapted isolates. Non-syncytium-inducing (NSI) primary isolates can be cultured only in the primary T cells from peripheral blood.
Early optimism for an AIDS vaccine was also engendered from studies on inactivated virus preparations of Simian Immunodeficiency Virus (SIV). Similarities between HIV-1 and SIV in morphology, genetic organization, infection and disease processes made SIV infection in rhesus monkeys an excellent model in which to explore different AIDS vaccine strategies. Early studies in this model showed that inactivated preparations of SIV grown on human T cell lines and formulated in adjuvant can protect macaques from infection after experimental inoculation with highly infectious, pathogenic variants of human cell-grown SIV (Desrosiers et al., PNAS USA, 1989, 86:6353-6357; Murphey-Corb et al., Science, 1989, 246:1293). Unexpectedly, this protection was lost when the SIV stock grown on homologous monkey cells was used for the challenge of immunized animals.
Later it was shown through immunization studies with monkey cell-grown SIV and uninfected human cells that protection from infection in those early SIV studies probably resulted from the stimulation of immune responses to xenogeneic human host cell proteins rather than to virus-encoded antigens (Stott, Nature, 1991, 353:393). Passive immunization experiments involving SIV have provided some evidence to suggest that certain anti-cell antibodies may contribute to protection against SIV infection in the absence of cell-mediated immunity (Gardner et al., AIDS Res Hum Retroviruses, 1995, 11:843-854).
The mechanism for the protective immunity to SIV challenge provided by anti-cell antibodies has not been delineated. One proposed mechanism is that the anti-cell mediated protection from SIV infection may involve virus-associated cell proteins. Immunodeficiency viruses such as HIV and SIV are known to incorporate cellular proteins probably obtained from the host cell as the viruses bud from the host cell membrane. Antibodies to some of the major histocompatibility complex (MHC) associated cellular proteins, .beta..sub.2 -microglobulin, HLA-DR, and HLA class I molecules, have been implicated in the in vitro neutralization of laboratory strains of SIV and HIV-1 (Arthur et al., Science, 1991, 258:1935). In addition to the MHC associated surface proteins, Montefiori et al. (Virology, 1994, 205:82; AIDS Res Hum Retroviruses, 1995, 11:1429) recently identified three complement control proteins, CD46, CD55, and CD59, on the surface of human cell-grown SIV and HIV-1 raising the possibility that these host cell molecules could also be utilized by the viruses as a mechanism to evade complement virolysis. An alternative proposed mechanism is that protection by anti-cell antibodies may be mediated by blocking the activity of immunodeficiency viruses against immune system cells in a previously unrecognized manner. There appears to be a host cell antigen complex associated with CD4 on the surface of the host T-cells which facilitates viral binding and entry and which may act as a target for protective anti-cell antibodies.
In addition to the CD4 receptor of the host antigen complex for binding HIV, other factors or HIV co-receptors affecting HIV replication, entry or fusion have recently been reported while the work on the present invention was on-going. These include three chemokines produced by CD8.sup.+ T cells (Cocchi et al., Science, 1995, 270:1811-1815) which are reported to be HIV suppressive; a co-receptor CXC-CKR4 (also termed fusin or LESTR) on CD4 expressing cells for T-tropic but not M-tropic HIV-1 fusion and entry (Feng et al., Science, 1996, 272:873), a .beta.-chemokine receptor CC-CKR5 which binds the three inhibitory chemokines (Cocchi et al.) as a co-receptor for M-tropic, but not T-tropic HIV-1 (Doranz et al., Cell, 1996, 85:1149; Dragic et al., Nature, 1996, 381:667; Choe et al. Cell, 1996, 85:1135; Deng et al., Nature, 1996, 381:661; Alkhatib et al., Science, 1996, 272:1955), and other .beta.-chemokine receptors (CC-CKR2b and CC-CKR3) as co-receptors for M- and dual-tropic HIV (Doranz et al., Cell, 1996, 85:1149). These co-receptors are disclosed to be previously described G protein coupled receptors with seven transmembrane segments (Loetscher et al., J Biol Chem, 1994, 264:232; Samson et al., Biochemistry, 1996, 35:3362).
To more precisely identify the putative cellular protein that may be stimulating protective responses, and to better characterize the mechanism of protection mediated by anti-cell antibodies, HIV-neutralizing activities were characterized by the present inventor for members of a panel of monoclonal antibodies directed against multiple cellular antigens: .beta..sub.2 microglobulin, MHC class I HLA A,B,C, MHC class II HLA DR proteins, and other T cell antigens associated with a T cell line, to identify those that are capable of neutralizing HIV primary isolates in an in vitro microplaque neutralization assay. Experiments were conducted to determine the scope of such neutralizing activity, if any, for the antibodies found to possess such activity; and to determine whether such in vitro activity can be translated into in vivo efficacy in an appropriate animal model(s).
The results of these experiments demonstrated that, except for antibodies directed against a host cell antigen complex comprising CD4, no other anti-cell antibodies including exclusively CD4-specific antibodies, can neutralize HIV-1 primary isolates as effectively in neutralization assays. The results also showed that antibodies directed against a host cell antigen complex comprising CD4 in association with domains from chemokine receptors exhibit enhanced binding with rsCD4. Further, antibodies with the desired properties are identified and can block in vivo SIV infection in monkeys and in vivo HIV-1 infection of the human immune system reconstituted in mice.